Figure 7. 80K-H is phosphorylated in vitro.

GST-tagged 80K-H or GST was coupled with glutathione beads and exposed to cell extracts of cos-1 cells transiently transfected with full-length HA-tagged PKCζ (kinase active) or HA-tagged PKCζ-CB (kinase-dead). After extensive washing, complexes were incubated in the presence of [γ-³²P]ATP. Samples were resolved by SDS/PAGE (10% polyacrylamide) and the gel was stained with Coomassie Brilliant Blue. The band corresponding to the GST construct was excised and incorporation of ³²P was determined by scintillation counting. Results are the means±S.E.M. for four independent experiments.